The voltage-sensitive sodium channel is an integral component of impulse conduction by neuronal cells, transmitting the initial inward current during an action potential. The channel has been purified from a number of different tissues, and consists of one large subunit termed alpha which is associated with one or two small subunits termed beta in certain tissues. in addition to the heterogeneity with respect to small subunits, the sodium channels from different tissues also express a variety of electrophysiological and toxin binding differences. Complementary DNA clones encoding the a subunit have been isolated from both rat brain and eel electroplax, and these have demonstrated that there are multiple different sodium channel a subunit genes expressed in rat brain. The primary goal of this proposal is to investigate the molecular basis for this diversity in voltage-sensitive sodium channel function. Differences with respect to the beta subunits will be examined by isolating cDNA clones encoding the two beta subunits from rat brain and studying their effects on sodium channel function following injection of in vitro transcribed RNA into Xenopus oocytes. The resulting channels will be characterized both biochemically and electrophysiologically. Tissue specific heterogeneity will be investigated by injecting RNA from different tissues into oocytes, and then characterizing the channels with respect to their tissue specific properties. The final approach involves directly examining functional sites within the sodium channel molecule by in vitro mutagenesis and using anti-peptide antibodies to perturb channel function. Ultimately, these studies should help to correlate molecular structure with the electrophysiological differences among voltage-sensitive sodium channels.